The collection, preservation and storage of tissue samples for molecular analysis is essential for cancer treatment and for research and development of tissue-based biomarkers for disease pathophysiology. Much effort is currently focused on determining markers based on nucleotides (i.e. DNA, mRNA, miRNA), proteins and metabolites for cancer staging, prognosis and treatment selection. Detection of these biomarkers would more efficiently direct patients to treatments with the highest potential for benefit.
Current tissue preservation methods such as formalin-fixed, paraffin embedded (FFPE) are suitable for histopathology studies but not amenable to biomarker analysis due to poor protein and nucleic acid recovery. Even though recent reports describe some success with genetic analysis, the poor quality of DNA and RNA restricts analysis by common techniques such as RT-PCR, microarrays and sequencing. Extraction of full-length, non-degraded, immunoreactive proteins from FFPE tissue has also proved challenging, with limited detection by common methods such as ELISA and bead-based multiplexed immunoassays.
Another tissue preservation method, snap-freezing of sectioned tissue samples in liquid nitrogen followed by storage at −80° C., has proved more successful for long-term tissue storage and protein and DNA analysis. While this method may be suitable for limited sampling, it is not practical for wide-scale use due to high costs and infrastructure requirements as well as logistical issues in collecting, maintaining and shipping samples at sub-freezing temperatures.
A major limitation for disease and biomarker research is a lack of robust and relevant biological samples. Small collections of biological samples are spread throughout research institutions, but sample collection and storage is not uniform and samples are often compromised, which can lead to faulty data. As a key recent example, NCI attempted to form a cancer biobank through the Cancer Genome Atlas program but found that up to 99% of stored blood and tissue samples were unacceptable for research.
The development of alternative methods for simplified tissue sample storage has proved challenging. Meanwhile, methods for blood specimen collection and storage are undergoing a revolution to a new method, known as dried blood spot (DBS) sampling, which offers considerable advantages over traditional venipuncture including decreased costs, reduced sample size and increased analyte stability. Using a finger or neonatal heel stick, approximately 100 μL of blood is spotted onto a filter paper and dried at ambient temperature. Once dried, analytes including DNA/RNA, proteins and small molecules are stable at ambient temperature or under refrigeration for years. We recently demonstrated that detection of miRNA levels were equivalent between wet and dried blood. Analytes are extracted from the paper with solvent and measured by traditional methods including LC-MS/MS, RT-PCR, microarray, ELISA, etc.
Similar to dried blood, suspensions of tumor cells dried on filter paper show RNA and DNA stability for at least six months and suitability for PCR-based analysis, suggesting that dried tissue may also provide long-term stability. It would therefore be desirable to have a system that would provide for the collection, preservation and long-term storage of biological tissue samples in order to enable wide range testing of the samples that are not possible with currently existing methodologies.